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PubMed2006

Systematic review: smoking cessation intervention strategies for adults and adults in special populations.

BACKGROUND: While smoking cessation interventions have been shown to work, questions remain about how to increase their efficacy. PURPOSE: To examine strategies for effective tobacco treatment in adults and special populations. DATA SOURCES: MEDLINE, Cumulative Index to Nursing and Applied Health (CINAHL), Cochrane Library, Cochrane Clinical Trials Register, Psychological Abstracts, and Sociological Abstracts (1 January 1980 to 10 June 2005). STUDY SELECTION: Systematic reviews; randomized, controlled trials; and observational studies. DATA EXTRACTION: Two reviewers independently abstracted data on study design, population, sample size, treatment, outcomes, and quality. DATA SYNTHESIS: Findings from systematic reviews were summarized and compared with findings from original research published beyond date ranges included in the reviews. Strength of evidence was used to assess the body of evidence. Our review included studies evaluating the efficacy of cessation strategies, such as self-help, counseling, single pharmaceutical agents, combined pharmacotherapies, and pharmacotherapies combined with psychological counseling. Research findings consistent with previous reviews show that self-help strategies alone are ineffective, but counseling and pharmacotherapy used either alone or in combination can improve rates of success with quit attempts. Two studies of self-help materials reported discrepancies across effects. Five studies provided mixed results for counseling interventions. Fourteen studies provided sufficient evidence of the efficacy of single pharmacotherapy, combined pharmacotherapy, and psychological interventions either with or without pharmacotherapy. Few studies focused on ways to reach or treat special populations. Three studies with hospitalized patients had findings consistent with a previous review showing no strong evidence that clinical diagnosis affected the likelihood of quitting. New evidence was insufficient to address the effectiveness of interventions for persons with coexisting psychiatric conditions and substance abuse problems. LIMITATIONS: Previous systematic reviews variably cover the range of issues we addressed. More recent studies do not fill all gaps, especially those for persons with coexisting disease. CONCLUSIONS: Although self-help strategies alone marginally affect quit rates, individual and combined pharmacotherapies and counseling either alone or in combination can significantly increase cessation. Using effective smoking treatments is strongly encouraged for all populations, especially those with high and heavy rates of smoking, such as psychiatric and substance abuse populations.

Adolescent↗
PubMed2002

Analysis of gene expression in the bovine blastocyst produced in vitro using suppression-subtractive hybridization.

Successful embryonic development is dependent on time and location-specific expression of appropriate genes. Unfortunately, information on stage-specific gene expression during early embryonic development in the bovine is lacking. In the present study, we compared gene expression between in vitro-produced Day 7-8 intact blastocysts (driver) and Day 9-10 hatched blastocysts (tester) using suppression-subtractive hybridization. Pools of 30 embryos for both driver and tester were used in the RNA extraction process. From limited amounts of starting material ( approximately 400 ng of total RNA), a reverse transcription-polymerase chain reaction (PCR) procedure was used to amplify the mRNA and generate sufficient cDNA to conduct suppression-subtractive hybridization. The subtracted cDNA products were cloned, and 126 cDNAs representing expressed mRNAs were isolated, sized, single-pass sequenced, and compared to known sequences in GenBank. Ninety-two clones provided sequence information for further analysis. Among these, 31 exhibited high homology to known genes. Three, 26S proteasomal ATPase (PSMC3), casein kinase 2 alpha subunit (CK2), and phosphoglycerate kinase (PGK) were selected and further characterized using real-time quantitative PCR to assess their differential expression in hatched blastocysts. Overall, a 1.3-, 1.6-, and 1.5-fold increase in expression level was observed in hatched blastocysts compared with intact blastocyst for PSMC3, CK2, and PGK, respectively. These results show that construction of subtracted cDNA libraries from small numbers of embryos is feasible and can provide information on gene expression patterns during preattachment embryogenesis.

Adenosine Triphosphatases↗
PubMed2003

In situ generation of carbenes: a general and versatile platform for organocatalytic living polymerization.

A metal-free, organocatalytic approach to living polymerization using N-heterocyclic carbenes as nucleophilic catalysts generated and used in situ in a single-pot process is detailed. The N-heterocyclic carbene catalyst platform is extremely versatile, as the nature of the substituents has a pronounced effect of catalyst stability and activity toward different substrates. The generation of imidazolium- and thiazaolium-based carbenes was accomplished from the reaction of the corresponding salts with the appropriate bases. This allowed the rapid screening of libraries of catalysts that provided a basic understanding of catalyst structure (sterics, electronics, etc.) with the polymerization rate, control, substrate, and range of molecular weights. The imidazole-based catalysts were significantly more active toward ROP than the thiazolium-based analogues. No appreciable differences between imidazol-2-ylidene and imidazolin-2-ylidene catalysts were observed. Less sterically demanding carbenes were found to be more active toward ring-opening polymerization (ROP) than their sterically encumbered analogues for lactone polymerization. These data prompted the investigation of ionic liquid as a precatalyst reservoir in a phase-transfer polymerization with an immiscible THF solution of monomer and initiator. In situ activation of the ionic liquid generates carbene that migrates to the organic phase effecting living ROP. Precatalyst (ionic liquid) regeneration terminates polymerization. This simple reaction/recycle protocol readily allows repetitive ROPs from the ionic liquid using commercially available materials.

Journal Article↗
PubMed1992

Isolation of DNA markers linked to a beet cyst nematode resistance locus in Beta patellaris and Beta procumbens.

In cultivated beet no useful level of resistance of the beet cyst nematode (BCN) Heterodera schachtii Schm. has been found, unlike the situation in wild species of the section Procumbentes. Stable introgression of resistance genes from the wild species into Beta vulgaris has not been achieved, but resistant monosomic additions (2n = 18 + 1), diploids of B. vulgaris with an extra alien chromosome carrying the resistance locus, have been obtained. Here we describe a new series of resistant monosomic fragment addition material of B. patellaris chromosome 1 (pat-1). We further describe the cloning of a single-copy DNA marker that specifically hybridizes with a monosomic addition fragment of approximately 8 Mb (AN5-90) carrying the BCN resistance locus. This marker and another fragment-specific, single-copy DNA marker probably flank the BCN locus on the addition fragment present in the AN5-203 material, which is approximately 19 Mb in size. Furthermore, several specific repetitive DNA markers have been isolated, one of which hybridizes to AN5-90 and also to DNA from a smaller DNA segment of Beta procumbens, present in line B883, carrying a BCN resistance locus introgressed into the B. vulgaris genome. This suggests that the specific repetitive marker is closely linked to the BCN locus.

Animals↗
PubMed1981

The DNA sequences of cloned complex satellite DNAs from Hawaiian Drosophila and their bearing on satellite DNA sequence conservation.

A class of restriction endonuclease fragments near 185 bp in length and comprising approximately 20% of the genomes of 3 species of Hawaiian Drosophila has been cloned using bacteriophage M13. The nucleotide sequences of 14 clones have been determined and the variation between clones has been found to be due to deletions and base changes. Analyses of uncloned material show that the cloning system itself does not introduce the variation. The variation of the basic repeat within and between species is high; 15% due to deletions and 10% due to base changes. The Drosophila data are similar in many respects to both the 23 bp calf satellite results (Pech et al., 1979 b) and those from sequence analyses of the 170 bp primate restriction fragments (Rubin et al., 1979; Donehower et al., 1980, Wu and Manuelidis, 1980). The intraspecies level of base changes and deletions in the calf satellite approaches 25% as does that in the human/African green monkey/baboon comparisons. The between species variation in the primate group is near 35%. Direct sequencing methods thus reveal a widespread sequence heterogeneity in both invertebrate and mammalian satellite systems of long or short repeat length. This heterogeneity does not support the strict sequence conservation implied by the "library" hypothesis, which claims a functional role in speciation for the rigid conservation of satellite DNA sequences (Fry and Salser, 1977). Furthermore the Drosophila and primate data reveal that satellite DNAs can change rapidly, though nonrandomly, at the nucleotide sequence level in a relatively closely knit group such as the Hawaiian species, as well as in more distantly related species from amongst the primates. We draw two major conclusions. There is no universal attribute of satellite DNA sequence per se, the only biological variable to date being the amount of satellite DNA and its effect in the germ line. Many aspects of satellite DNA evolution conform to Kimura's (1979) concepts of neutrality.

Animals↗
PubMed1990

Down-regulation of a calmodulin-related gene during transformation of human mammary epithelial cells.

A human cDNA library obtained from cultured normal mammary epithelial cells (HMECs) was searched by subtractive hybridization for genes whose decrease in expression might be relevant to epithelial transformation. One clone identified by this procedure corresponded to a 1.4-kilobase mRNA, designated NB-1, whose expression was decreased greater than 50-fold in HMECs tumorigenically transformed in vitro after exposure to benzo[a]pyrene and Kirsten sarcoma virus. Sequence analysis of NB-1 cDNA revealed an open reading frame with a high degree of homology to calmodulin. NB-1 expression could be demonstrated by polymerase chain reaction amplification in normal breast, prostate, cervix, and epidermal tissues. The presence of NB-1 transcripts was variable in primary breast carcinoma tissues and undetectable in tumor-derived cell lines of breast, prostate, or other origins. NB-1 mRNA expression could be down-regulated in cultured HMECs by exposure to reconstituted extracellular matrix material, while exposure to transforming growth factor type beta increased its relative abundance. The protein encoded by NB-1 may have Ca2+ binding properties and perform functions similar to those of authentic calmodulin. Its possible roles in differentiation and/or suppression of tumorigenicity in epithelial tissues remain to be examined.

Base Sequence↗
PubMed2005

Microgenomics: gene expression analysis at the tissue-specific and single-cell levels.

Plant organs are composed of many different cell types and the analysis of 'bulk' material results in the average of all information in these cells. Therefore, this does not reflect any individuality of the tissues present in plants. This review briefly summarizes different sampling methods which provide tissue- and cell-specific samples, respectively. In addition, gene expression analysis tools that allow the analysis of transcripts in minute samples are discussed in detail. The combination of both approaches results in high resolution gene expression data, which increases understanding of plant physiology in such diverse areas as primary and secondary metabolism, plant defence or stress response.

Base Sequence↗
PubMed2001

Intravenous oxytocin alone for cervical ripening and induction of labour.

BACKGROUND: Oxytocin is the commonest induction agent used worldwide. It has been used alone, in combination with amniotomy or following cervical ripening with other pharmacological or non-pharmacological methods. Prior to the introduction of prostaglandin agents oxytocin was used as a cervical ripening agent as well. In developed countries oxytocin alone is more commonly used in the presence of ruptured membranes whether spontaneous or artificial. In developing countries where the incidence of HIV is high, delaying amniotomy in labour reduces vertical transmission rates and hence the use of oxytocin with intact membranes warrants further investigation. This review will address the use of oxytocin alone for induction of labour. Amniotomy alone or oxytocin with amniotomy for induction of labour has been reviewed elsewhere in the Cochrane Library. Trials which consider concomitant administration of oxytocin and amniotomy will not be considered. This is one of a series of reviews of methods of cervical ripening and labour induction using a standardised methodology. OBJECTIVES: To determine the effects of oxytocin alone for third trimester cervical ripening or induction of labour in comparison with other methods of induction of labour or placebo/no treatment. SEARCH STRATEGY: The Cochrane Pregnancy and Childbirth Group Trials Register, the Cochrane Controlled Trials Register and bibliographies of relevant papers. Last searched: May 2001. SELECTION CRITERIA: The criteria for inclusion included the following: (1) clinical trials comparing vaginal prostaglandins used for third trimester cervical ripening or labour induction with placebo/no treatment or other methods listed above it on a predefined list of labour induction methods; (2) random allocation to the treatment or control group; (3) adequate allocation concealment; (4) violations of allocated management not sufficient to materially affect conclusions; (5) clinically meaningful outcome measures reported; (6) data available for analysis according to the random allocation; (7) missing data insufficient to materially affect the conclusions. DATA COLLECTION AND ANALYSIS: A strategy was developed to deal with the large volume and complexity of trial data relating to labour induction. This involved a two-stage method of data extraction. The initial data extraction was done centrally, and incorporated into a series of primary reviews arranged by methods of induction of labour, following a standardised methodology. The data is to be extracted from the primary reviews into a series of secondary reviews, arranged by category of woman. MAIN RESULTS: In total, 110 trials were considered; 52 have been excluded and 58 included examining a total of 11,129 women. Comparing oxytocin alone with expectant management: Oxytocin alone reduced the rate of unsuccessful vaginal delivery within 24 hours when compared with expectant management (8.3% versus 54%, relative risk (RR) 0.16, 95% confidence interval (CI) 0.10,0.25) but the caesarean section rate was increased (10.4% versus 8.9%, RR 1.17, 95% CI 1.01,1.36). This increase in caesarean section rate was not apparent in the subgroup analyses. Women were less likely to be unsatisfied with induction rather than expectant management, in the one trial reporting this outcome (5.5% versus 13.7%, RR 0.43, 95% CI 0.33, 0.56). Comparing oxytocin alone with vaginal prostaglandins: Oxytocin alone was associated with an increase in unsuccessful vaginal delivery within 24 hours (52% versus 28%, RR 1.85, 95% CI 1.41, 2.43), irrespective of membrane status, but there was no difference in caesarean section rates (11.4% versus 10%, RR 1.12, 95% CI 0.95, 1.33). Comparing oxytocin alone with intracervical prostaglandins: Oxytocin alone was associated with an increase in unsuccessful vaginal delivery within 24 hours when compared with intracervical PGE2 (51% versus 35%, RR 1.49, 95% CI 1.12,1.99). For all women with an unfavourable cervix regardless of membrane status, the caesarean section rates were increased (19.0% versus 13.1%, RR 1.42, 95% CI 1.11, 1.82). REVIEWER'S CONCLUSIONS: Overall, comparison of oxytocin alone with either intravaginal or intracervical PGE2 reveals that the prostaglandin agents probably overall have more benefits than oxytocin alone. The amount of information relating to specific clinical subgroups is limited, especially with respect to women with intact membranes. Comparison of oxytocin alone to vaginal PGE2 in women with ruptured membranes reveals that both interventions are probably equally efficacious with each having some advantages and disadvantages over the others. With respect to current practice in women with ruptured membranes induction can be recommended by either method and in women with intact membranes there is insufficient information to make firm recommendations.

Cervical Ripening↗
PubMedDate not supplied

[Retroviral reporter systems for the assessment of activity of stress-induced signal transduction pathways controlled by p53, HIF-1 and HSF-1 transcription factors].

Tumor suppressor p53, hypoxia-inducible factor 1 (HIF-1) and heat-shock factor 1 (HSF-1) are involved as the key transcription factors in cellular response to stress, induced by genetic material damage, hypoxia and heat shock respectively. The protein factors listed above also play an integral part in tumor development and progression. Thus, modulation of their activity may be important for treatment of cancer. In our work we obtained the reporter constructs for quantitative assessment of p53, HIF-1 and HSF-1 transcriptional activity on the basis of retro- and lentiviruses, allowing to obtain reporter cell lines almost out of any cell type. Induction of beta-galactosidase reporter gene expression, reflecting the activity of p53 and HIF-1 factors, depends on dose of treatment and also correlates with the induction of the endogenous target genes expression. The observed effect of activating treatments completely disappeared when the expression of p53 and HIF-1 genes was inhibited with specific siRNAs. The obtained reporter constructs may find the application in the screening of chemical and genetic (such as siRNA- and cDNA-libraries) modulators of transcriptional activity along with the investigation of components of signal transduction pathways modulating the transcriptional activity of those factors.

Base Sequence↗
PubMed1993

Interactive videodisc and compact disc-interactive for ophthalmic basic science and continuing medical education.

BACKGROUND: The authors designed and implemented a complete curriculum in ophthalmic pathology using IBM- and Macintosh-based interactive videodisc (IVD) technology. They also redesigned a portion of this curriculum for a new television-based platform, compact disc-interactive (CD-I). METHODS: The following issues were addressed: curriculum design, instructional design, the assembly of illustrations and the ownership of such materials, the generation of computer-based medical art and animation, and programming. The issue of academic credit for faculty participating in this effort also was considered. RESULTS: The computer-based IVD program provides the following features: (1) rapid access to thousands of high-quality illustrations with the option of superimposing graphic labels and text directly over pictures; (2) the ability to view enlargements of photographs; (3) an online glossary to view definition of terms coupled with high-quality photographs; and (4) a dynamic introduction to pathophysiology using interactive animation sequences. The authors were able to incorporate the same interactive features into the CD-I version. High-quality medical illustrations can be used effectively on the CD-I platform. CONCLUSIONS: Computer-based multimedia workstations are relatively expensive for personal use but may be useful if the equipment can be shared in a learning center or library. Compared with interactive computer-based solutions, consumer-oriented television-based technology such as CD-I is a relatively inexpensive vehicle for providing continuing medical education programs intended for use in the individual practitioner's office or home.

Compact Disks↗
PubMed1996

SCO-spondin: a new member of the thrombospondin family secreted by the subcommissural organ is a candidate in the modulation of neuronal aggregation.

A number of cues are known to influence neuronal development including growth factors, cell-adhesion molecules, components of the extracellular matrix and guidance molecules. In this study, we present molecular and functional evidence that SCO-spondin, a novel relative of the thrombospondin family, could also be involved in neuronal development by modulating cell aggregative mechanisms. SCO-spondin corresponds to glycoproteins secreted by the subcommissural organ (SCO), an ependymal differentiation of the vertebrate brain located at the entrance to the Sylvian aqueduct. A cDNA clone of 2.6 kb, isolated from a bovine SCO cDNA library, was shown to be specifically and highly expressed in the bovine SCO by in situ hybridization and was subsequently sequenced. Analysis of the deduced amino acid sequence reveals the presence of four conserved domains known as thrombospondin (TSP) type I repeats. To account for the homology with thrombospondins and F-spondin, this secreted glycoprotein was called SCO-spondin. Two potent binding sites to glycosaminoglycan (BBXB) and to cytokine (TXWSXWS) are also found in the TSP type I repeats. The deduced amino acid sequence exhibits three other conserved domains called low density lipoprotein (LDL) receptor type A repeats. The possibility of SCO-spondin involvement in neuronal development as a component of the extracellular matrix is discussed regarding these molecular features. The idea of a modulation of cell-cell and/or cell-matrix interaction is further supported by the anti-aggregative effect observed on cultured neuronal cells of material solubilized from Reissner's fiber. That Reissner's fiber, the condensed secretory product of the SCO present along the whole spinal cord can be a potent morphogenetical structure is an important concept for the analysis of the molecular mechanisms leading to spinal cord differentiation.

Amino Acid Sequence↗
PubMed1991

30-kDa heparin-binding protein of brain (amphoterin) involved in neurite outgrowth. Amino acid sequence and localization in the filopodia of the advancing plasma membrane.

A cDNA library constructed from mRNA of rat brain was used to clone the cDNA that encodes the 30-kDa heparin-binding protein (amphoterin) that is developmentally regulated in brain and enhances neurite outgrowth in cerebral neurons. cDNA and peptide sequencing identified a dipolar sequence that has been previously found in studies of high mobility group 1 protein: the 184-amino acid cationic region is followed by a cluster of 30 anionic residues. The mRNA encoding amphoterin is also developmentally regulated; it is strongly reduced in quantity after the rapid perinatal growth phase of the rat brain. Anti-synthetic peptide antibodies raised according to the sequence of amphoterin were shown to bind specifically to the protein isolated from brain, and were used to detect amphoterin in subcellular fractions and in immunostaining of cells. Amphoterin was found in the cytoplasm of the cell soma, in the cell processes, and the substrate-attached material. In cells that are at an active stage of spreading and extending their cytoplasmic processes amphoterin was especially associated with plasma membrane filopodia. The distinct localization to the filopodia of the advancing plasma membrane suggests that endogenous amphoterin has a role in the extension of neurite-type cytoplasmic processes in developing cells. This inference is further supported by the finding that both anti-amphoterin and the anti-synthetic peptide antibodies in the culture media strongly inhibit the outgrowth of cytoplasmic processes.

Amino Acid Sequence↗
PubMed1995

Molecular genetics and the diagnostic mycology laboratory.

The laboratory diagnoses or confirmation of fungal diseases are extremely important considerations in the proper and efficient management of patients experiencing these infections. When an infectious process is suspected, suitable material must be collected and submitted to the pathologist for microscopic examination and to the diagnostic laboratory for culture. The practice of modern medicine dictates that the turn-around time be reduced to a minimum so that specific therapy can be rapidly instituted. Unfortunately, the classical procedures used in processing specimens for isolation and identification of fungi implicated in these diseases are technically time consuming and on occasion labor-intensive. More importantly, fungi are slow growing and cultures are frequently held for 3 to 4 weeks before they are discarded as negative. With the exception of coccidioidomycosis, most attempts to develop serological tests for rapid diagnosis of fungal infections have been hampered by poor specificity caused by immunologic cross reactivity. To some extent, such problems can be circumvented by application of molecular biological techniques to these problems. For example, procedures have been developed that allow efficient screening of DNA expression libraries to identify useful recombinant antigens and production of selected antigens in quantity. This is potentially a practical approach using molecular biological techniques for development of sensitive, specific and practical reagents useful in the serological diagnosis of fungal infections. In addition, nucleic acid hybridization techniques based on the ability of complementary nucleic acid strands to specifically align and associate to form stable double-stranded complexes have been put to use to develop confirmation tests for a variety of important fungal isolates.(ABSTRACT TRUNCATED AT 250 WORDS)

Antigens, Fungal↗
PubMed1993

Cloning, sequencing, and expression of a cDNA encoding rat liver carnitine palmitoyltransferase I. Direct evidence that a single polypeptide is involved in inhibitor interaction and catalytic function.

We report the isolation and characterization of a full-length cDNA encoding rat liver carnitine palmitoyltransferase I (CPT I). Oligonucleotides corresponding to two tryptic peptides derived from the malonyl-CoA/etomoxir-CoA-binding protein of rat liver mitochondria (Esser, V., Kuwajima, M., Britton, C. H., Krishnan, K., Foster, D. W., and McGarry, J. D. (1993) J. Biol. Chem. 268, 5810-5816) were used to screen a rat liver cDNA library constructed in the plasmid cloning vector, pcDV. The clone obtained consisted of a 102-nucleotide 5'-untranslated region, a single open reading frame of 2,319 bases predicting a protein of 773 amino acids (M(r) = 88,150), and a 3'-untranslated segment of 1,957 nucleotides followed by the poly(A)+ tail. A 0.9-kilobase fragment of the cDNA recognized a single species of mRNA (approximately 4.7 kilobases in size) in rat liver. The identity of the cDNA was confirmed by the findings that (i) the open reading frame encoded all four peptides found in the original protein; (ii) transfection of COS cells with the cDNA subcloned into the expression vector, pCMV6, resulted in a selective and 10-20-fold induction of a malonyl-CoA- and etomoxir-CoA-sensitive CPT activity; and (iii) the overexpressed product was readily detected on Western blots by an antibody raised against the starting material. It seems likely that the de novo synthesized enzyme is targeted to the mitochondrial outer membrane via a leader peptide and that the mature protein achieves membrane anchoring through a stretch of 20 amino acids present near its amino terminus. The predicted amino acid sequence of the protein shows regions of strong identity with those of three other rat acyltransferases, namely, liver CPT II, liver carnitine octanoyltransferase, and brain choline acetyltransferase. The findings provide the first insight into the structure of a CPT I isoform. They also establish unequivocally that CPT I and CPT II are distinct proteins and that inhibitors of CPT I interact within its catalytic domain, not with an associated regulatory component.

Amino Acid Sequence↗
PubMed1998

Fasciola hepatica: characterization and cloning of the major cathepsin B protease secreted by newly excysted juvenile liver fluke.

Proteolytic activity present in the excreted/secreted (ES) material of newly excysted juvenile (NEJ) Fasciola hepatica was biochemically analyzed. By gelatin substrate SDS-PAGE, only one region of activity was observed in the NEJ ES material at a molecular mass of 29 kDa. Both the secreted cathepsin L from adult fluke and the 29-kDa proteolytic activity of NEJ ES show a common pH optimum of 7.5, a cysteine protease inhibition profile, and preference for the N-benzyloxycarbonyl (Z)-Phe-Arg-NHMec fluorogenic substrate over Z-Arg-Arg-NHMec and Z-Arg-NHMec. In vitro analysis revealed that the NEJ protease activity digested sheep immunoglobulin heavy chain and bovine serum albumin but not bovine hemoglobin. Amino-terminal protein sequence analysis of the 29-kDa NEJ protease band revealed two sequences with homology to the cathepsin B family of proteases. Using degenerate oligonucleotides designed from the N-terminal sequence, reverse transcriptase polymerase chain reaction with NEJ RNA amplified a cDNA sequence encoding the first 236 amino acids of mature cathepsin B. Using this cDNA fragment an overlapping cDNA was isolated from a LambadaZAP cDNA library constructed with poly(A)+ RNA from immature 5-week-old liver fluke. Together with the N-terminal sequence, these cDNAs predict a mature cathepsin B sequence of 254 amino acids which shows 48-51% sequence identity to mammalian and Schistosoma mansoni cathepsin B. We conclude that, in contrast to the major proteases released by adult fluke, the major secreted protease of NEJ of F. hepatica is of the cathepsin B class.

Amino Acid Sequence↗
PubMed2006

Probe hybridization array typing: a binary typing method for Escherichia coli.

The ability to distinguish between Escherichia coli strains is critical for outbreak investigations. Binary typing, based on the presence or absence of genetic material, provides a high-throughput alternative to gel- and PCR-based typing techniques that generate complex banding patterns and lack uniform interpretation criteria. We developed, validated, and determined the discriminatory power of an E. coli binary typing method, probe hybridization array typing (PHAT). In PHAT, the absence or presence of genetic material is identified by using DNA hybridization to produce a reproducible and portable fingerprint for each genome. PHAT probes were generated from genome subtractive hybridization experiments. We PHAT typed the ECOR collection of strains from a variety of geographical locations, and 33 rectal E. coli strains selected from college-aged women with urinary tract infection. In the set of 33 human rectal strains, the discriminatory power of PHAT (98%) equaled that of multilocus sequence typing (MLST) and pulsed-field gel electrophoresis. However, for ECOR strains, which include nonhuman strains, the current set of PHAT probes was less discriminating than MLST, ribotyping, and enterobacterial repetitive intergenic consensus sequence PCR (80% versus 97, 92, and 97%, respectively). When we limited the analysis to ECOR strains of B2 and D lineage, which are associated with human infection, current PHAT probes were highly discriminatory (94%). PHAT can be applied in a high-throughput format (i.e., "library on a slide"), the discriminatory ability can be varied based on the probe set, and PHAT is readily adapted to other bacterial species with high variation in genetic content.

Adolescent↗
PubMed1995

Balancing learner control and realism with specific instructional goals: case studies in fluid balance for nursing students.

In designing instruction, there is an inherent tension between permitting learner control and focusing on specific instructional goals. This tension is particularly problematic in computer-based clinical case studies. In addition, the question of the appropriate level of realistic detail is central to case study design. More learner control and greater detail require greater complexity and interactivity, and consequently longer development time per case. They may also lead to difficulty in focusing on specific content. This program is designed to present computer-based case studies as an adjunct to classroom instruction in a specific content area, with significant compromises in realism compensated by the potential to focus on a specific set of problems and expose students to a fairly large number of "patients." The degree of learner control is necessarily limited by the program's structure, but the design attempts to allow a considerable amount of self-direction within those limits. The program's main path goes through a series of case studies illustrating a variety of fluid and electrolyte problems. These case studies are fairly simple and tightly structured: history, physical assessment, and laboratory data are presented near the beginning of each study, and learners are given predefined choices with immediate feedback at specified decision points. The case studies are designed to be completed fairly rapidly, typically no more than about thirty minutes per case. The distortion of the case study format by the obvious focus on fluid and electrolyte problems is compensated somewhat by inclusion of a number of cases with no relevant problems. Within this structured format, the program gives the learner control over a number of aspects of the learning experience. Hypertext is used hierarchically, with two levels of explanation for some central concepts and one level for material not directly relevant to the main instructional goals. Consequently, the learner controls content density to a considerable degree. Since nursing students are under pressure to absorb large amounts of clinically relevant information within a relatively short time, the availability of additional information on diagnoses, charting abbreviations, normal ranges for lab values, etc., may provide an added incentive to use the program. Though the case studies are linear with no branching, learners have several options in using them. The basic user mode involves simply going through the case studies and "solving" them: evaluating the data, identifying problems, and making basic treatment decisions. Alternatively, the learner may choose a particular problem and use the program as a "semi-tutorial." Backward movement is allowed, either to review the development of a case or to try again after an incorrect response. Accumulated information is available in an on-screen "chart" for ready reference. A "reference library" is accessible for browsing from any point in the program. Learner evaluation of the program is in progress.

Computer Simulation↗
PubMed1990

Vision assisted robotics and tape technology in the life-science laboratory: applications to genome analysis.

Recent proposals to analyze a number of major genomes have created a need for methods of rapidly manipulating and assaying very large numbers of specimens containing DNA fragments. Systems for sampling semi-solid biological material from mixed disordered arrays are required. These samples need to be sorted in an ordered format and stored in minimum space at known locations. It should be possible to recall them quickly from stock, individually or collectively, for duplication or re-ordering into new arrays for appropriate analysis. This article reviews a range of problems associated with high speed multiple specimen handling and assay in the molecular biology laboratory and outlines the solutions currently available to address these difficulties. Designs for automatic systems to manipulate biosamples and analyze DNA are presented.

Biotechnology↗